College Forensics: Project Advance Chapter 12: Forensic Serology

What is this chapter about? Blood types and their antigens and antibodies. Blood types and their antigens and antibodies. Agglutination. Agglutination. Whole blood typing. Whole blood typing. Characterizing a stain as blood. Characterizing a stain as blood. Significance of the precipitin test in serology. Significance of the precipitin test in serology. Absorption-elution techniques. Absorption-elution techniques. Contribution of polymorphic enzymes and proteins to blood’s individualization. Contribution of polymorphic enzymes and proteins to blood’s individualization. Chromosomes and genes. Chromosomes and genes. Significance of Punnett square. Significance of Punnett square. Lab tests for seminal stains. Lab tests for seminal stains. Preservation of stains for lab examination. Preservation of stains for lab examination. Collection of physical evidence in rape investigation. Collection of physical evidence in rape investigation.

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DNA: abbreviation for deoxyribonucleic acid- the molecules carrying the body’s genetic information. DNA is double- stranded in the shape of a double helix. DNA: abbreviation for deoxyribonucleic acid- the molecules carrying the body’s genetic information. DNA is double- stranded in the shape of a double helix. Plasma: the fluid portion of unclotted blood. Plasma: the fluid portion of unclotted blood. Erythrocyte: a red blood cell. Erythrocyte: a red blood cell. Serum: the liquid that separates from the blood when a clot is formed. Serum: the liquid that separates from the blood when a clot is formed. Antigen: a substance, usually a protein that stimulates the body to produce antibodies against it. Antigen: a substance, usually a protein that stimulates the body to produce antibodies against it. Antibody: a protein that destroys or inactivates a specific antigen. Antibodies are found in the blood serum. Antibody: a protein that destroys or inactivates a specific antigen. Antibodies are found in the blood serum. Antiserum: blood serum in which there are specific antibodies. Antiserum: blood serum in which there are specific antibodies. Agglutination: the clumping together of red blood cells by the action of an antibody. Agglutination: the clumping together of red blood cells by the action of an antibody. Terms to Know:

Serology: the study of antigen-antibody reactions. Serology: the study of antigen-antibody reactions. Hemoglobin: a red blood cell protein responsible for transporting oxygen in the bloodstream and the red coloring of blood. Hemoglobin: a red blood cell protein responsible for transporting oxygen in the bloodstream and the red coloring of blood. Precipitin: an antibody that reacts with its corresponding antigen to form a precipitate. Precipitin: an antibody that reacts with its corresponding antigen to form a precipitate. Secretor: an individual who secretes his or her blood-type antigen(s) in body fluids. Approximately 80 percent of the population are secretors. Secretor: an individual who secretes his or her blood-type antigen(s) in body fluids. Approximately 80 percent of the population are secretors. Enzyme: a type of protein that acts as a catalyst for certain specific reactions. Enzyme: a type of protein that acts as a catalyst for certain specific reactions. Polymorphism: the existence of more than one form of a genetic trait. Polymorphism: the existence of more than one form of a genetic trait.

Iso-enzymes: multiple molecular forms of an enzyme, each having the same or very similar enzyme activities. Iso-enzymes: multiple molecular forms of an enzyme, each having the same or very similar enzyme activities. Gene: a unit of inheritance consisting of a DNA segment located on a chromosome. Gene: a unit of inheritance consisting of a DNA segment located on a chromosome. Chromosome: a rodlike structure in the cell nucleus, along witch the genes are located. It is composed of DNA surrounded by other material, mainly proteins. Chromosome: a rodlike structure in the cell nucleus, along witch the genes are located. It is composed of DNA surrounded by other material, mainly proteins. Egg: the female reproductive cell. Egg: the female reproductive cell. Sperm: the male reproductive cell. Sperm: the male reproductive cell. Zygote: the cell arising from the union of an egg and a sperm cell. Zygote: the cell arising from the union of an egg and a sperm cell. X-chromosome: the female sex chromosome. X-chromosome: the female sex chromosome. Y-chromosome: the male sex chromosome. Y-chromosome: the male sex chromosome.

Locus: the physical location of a gene on a chromosome. Locus: the physical location of a gene on a chromosome. Allele: any of several alternative forms of a gene located at the same point on a particular pair of chromosomes. For example, the genes determining the blood types A and B are alleles. Allele: any of several alternative forms of a gene located at the same point on a particular pair of chromosomes. For example, the genes determining the blood types A and B are alleles. Homozygous: having two identical allelic genes on two corresponding positions of a pair of chromosomes. Homozygous: having two identical allelic genes on two corresponding positions of a pair of chromosomes. Heterozygous: having two different allelic genes on two corresponding positions of a pair of chromosomes. Heterozygous: having two different allelic genes on two corresponding positions of a pair of chromosomes. Genotype: the particular combination of genes present in the cells of an individual. Genotype: the particular combination of genes present in the cells of an individual. Phenotype: the physical manifestation of a genetic trait such as shape, color, and blood type. Phenotype: the physical manifestation of a genetic trait such as shape, color, and blood type.

Acid Phosphatase: an enzyme found in high concentration in semen. Acid Phosphatase: an enzyme found in high concentration in semen. Oligospermia: a condition describing an abnormally low sperm count. Oligospermia: a condition describing an abnormally low sperm count. Aspermia: the absence of sperm; sterility in males. Aspermia: the absence of sperm; sterility in males.

Karl Landsteiner In 1901, recognized that not all human blood was the same. For many years, blood transfusions often ended in failure. In 1901, recognized that not all human blood was the same. For many years, blood transfusions often ended in failure. Landsteiner’s discovery led to the classification system called the A-B-O system, allowing doctors to properly match donor and recipient blood types. Landsteiner’s discovery led to the classification system called the A-B-O system, allowing doctors to properly match donor and recipient blood types. By 1937, the Rh factor had been discovered, and now more than 100 blood factors have been shown to exist. By 1937, the Rh factor had been discovered, and now more than 100 blood factors have been shown to exist. Until the 1990s, criminalists focused on blood factors as a way to individualize crime scene blood. Until the 1990s, criminalists focused on blood factors as a way to individualize crime scene blood. DNA technology has since become the predominant method of individualization with blood samples. DNA technology has since become the predominant method of individualization with blood samples.

Blood A complex mixture of cells, enzymes, proteins & inorganic substances A complex mixture of cells, enzymes, proteins & inorganic substances Fluid portion of blood is called the plasma (55% of blood content): Fluid portion of blood is called the plasma (55% of blood content): Primarily water Primarily water Red cells (erythrocytes) Red cells (erythrocytes) White cells (leukocytes) White cells (leukocytes) Platelets (thrombocytes) Platelets (thrombocytes)

What is Serology? A term which describes laboratory tests which employ a specific antigen and serum antibody reactions. A term which describes laboratory tests which employ a specific antigen and serum antibody reactions. Blood clots when a protein in the plasma known as fibrin traps and enmeshes RBCs. Blood clots when a protein in the plasma known as fibrin traps and enmeshes RBCs. If one were to remove the clotted material, a pale yellowish liquid known as serum would be left. If one were to remove the clotted material, a pale yellowish liquid known as serum would be left.

Erythrocytes Transport oxygen from the lungs to the body tissues Transport oxygen from the lungs to the body tissues Transport carbon dioxide from the tissues to the lungs Transport carbon dioxide from the tissues to the lungs Red cells possess chemical structures on their surfaces called antigens or agglutinogens: Red cells possess chemical structures on their surfaces called antigens or agglutinogens: Impart blood type characteristics: Impart blood type characteristics: A antigens A antigens B antigens B antigens D antigens (Rh factor) D antigens (Rh factor)

Blood Type Three types (alleles) of blood type gene: Three types (alleles) of blood type gene: I A, I B, i I A, I B, i Each individual inherits one blood type gene from their mother & one from their father Each individual inherits one blood type gene from their mother & one from their father 6 possible combinations (genotypes) 6 possible combinations (genotypes) I A I A, I A i, I B I B, I B i, I A I B, ii I A I A, I A i, I B I B, I B i, I A I B, ii Genotype determines blood type Genotype determines blood type

Antibodies or Agglutinins Proteins that are present in the serum, called antiserum. Proteins that are present in the serum, called antiserum. Responsible for ensuring that the only blood cells that can survive in a person are cells of the correct blood type. Responsible for ensuring that the only blood cells that can survive in a person are cells of the correct blood type. Agglutination: the clumping together of RBCs by the action of an antibody. Agglutination: the clumping together of RBCs by the action of an antibody. Antibodies produced by the A alleles remove any red blood cells containing B antigens by clumping them together. Antibodies produced by the A alleles remove any red blood cells containing B antigens by clumping them together. Antibodies produced by the B alleles remove any red blood cells possessing A antigens. Antibodies produced by the B alleles remove any red blood cells possessing A antigens.

Type O Blood Possessed by people whose genotype is ii Possessed by people whose genotype is ii Both parents passed on the i gene Both parents passed on the i gene Have no antigens Have no antigens These cells can be introduced into a person with Type A or Type B because these cells are not attacked by the antibodies these people possess These cells can be introduced into a person with Type A or Type B because these cells are not attacked by the antibodies these people possess Have both a & b antibodies Have both a & b antibodies Can only have other O type cells mixed with this blood Can only have other O type cells mixed with this blood

Type A Possessed by people with genotype I A I A or I A i Possessed by people with genotype I A I A or I A i I A is dominant to i I A is dominant to i Possesses antibody b Possesses antibody b will destroy any Type B red cells will destroy any Type B red cells compatible with A or O red cells compatible with A or O red cells What’s the difference between homozygous and heterozygous? What’s the difference between homozygous and heterozygous?

Type B Possessed by people with genotype I B I B or I B i Possessed by people with genotype I B I B or I B i I B is dominant to i I B is dominant to i Possesses antibody a Possesses antibody a will destroy any Type A red cells will destroy any Type A red cells compatible with B or O red cells compatible with B or O red cells

Type AB Possessed by people with genotype I A I B Possessed by people with genotype I A I B I A and I B are codominant I A and I B are codominant Possesses no antibodies Possesses no antibodies can have A, B, AB, or O cells added can have A, B, AB, or O cells added can’t be added to any other blood type without being destroyed by an antibody can’t be added to any other blood type without being destroyed by an antibody

Antibodies are normally bivalent, having two reactive sites

Blood Typing Blood typing is done by reacting whole blood with antibody A and antibody B, and then observing the reactions. Blood typing is done by reacting whole blood with antibody A and antibody B, and then observing the reactions.

Blood Typing

Relative Frequency of Blood Types in Human Populations

Immunoassay : Immunoassay: A technique utilizing antibodies to bind specifically to targeted substances in a specimen in order to identify their presence. Used to detect drugs in blood and urine. Antibody Antibody - Protein produced by the immune system of humans and higher animals in response to the presence of a specific antigen. Antigen Antigen - Substance that can trigger an immune response, resulting in production of an antibody as part of the body's defense against infection and disease. Many antigens are foreign proteins (those not found naturally in the body). An allergen is a special type of antigen which causes an antibody response.

Immunoassay Antibodies that react with drugs do not naturally exist. Antibodies that react with drugs do not naturally exist. They can be produced in animals such as rabbits by first combining the drug with a protein and injecting this combination into the animal. They can be produced in animals such as rabbits by first combining the drug with a protein and injecting this combination into the animal. This drug-protein complex acts as an antigen stimulating the animal to produce antibodies. This drug-protein complex acts as an antigen stimulating the animal to produce antibodies. The recovered blood serum of the animal will contain antibodies that are specific to the drug. The recovered blood serum of the animal will contain antibodies that are specific to the drug.

Immunoassay Immunoassay: A technique utilizing antibodies to bind specifically to targeted substances in a specimen in order to identify their presence. EMIT - Enzyme Multiplied Immunoassay Technique A competitive binding immunoassay that avoids the separation step. As Test antigen is increased it occupies more of the antibody molecules, so fewer of the enzyme labels are blocked. Substrate → product response increases. TWO METHODS

Immunoassay Immunoassay: A technique utilizing antibodies to bind specifically to targeted substances in a specimen in order to identify their presence. ELISA - Enzyme Linked Immunosorbent Assay Wide application - diagnosis & research. Non-competitive "sandwich" form requires two antibody- recognition sites on test antigen. TWO METHODS

Forensic Characterization of Dried Bloodstains Is it blood? Is it blood? From what species? From what species? If human, how closely can it be associated with a particular individual? If human, how closely can it be associated with a particular individual?

Determination of Blood The Kastle-Meyer color test uses the chemical phenolphtalein The Kastle-Meyer color test uses the chemical phenolphtalein When a bloodstain, phenolphtalein reagent, and hydrogen peroxide are mixed, hemoglobin causes the formation of a deep pink color When a bloodstain, phenolphtalein reagent, and hydrogen peroxide are mixed, hemoglobin causes the formation of a deep pink color Hemastix strips, moistened with distilled water, placed in contact with bloodstain → green color indicates blood Hemastix strips, moistened with distilled water, placed in contact with bloodstain → green color indicates blood Luminol produces light when sprayed on blood sample Luminol produces light when sprayed on blood sample Is it human? Precipitin testing is performed Is it human? Precipitin testing is performed Human antiserum is obtained from rabbits Individualization: now focuses on DNA profiling, rather than examining the uniqueness of proteins found in a person’s blood

Forensic Characterization of Semen Acid Phosphatase test: useful for detecting seminal stains Acid Phosphatase test: useful for detecting seminal stains Acid Phosphatase is an enzyme secreted by the prostate gland, and can easily be detected when it comes in contact with an acidic solution of sodium alpha naphthylphosphate and Fast Blue B dye. A purple color is a positive indication. Acid Phosphatase is an enzyme secreted by the prostate gland, and can easily be detected when it comes in contact with an acidic solution of sodium alpha naphthylphosphate and Fast Blue B dye. A purple color is a positive indication. Also, 4-methyl umbelliferyl phosphate (MUP) fluoresces under UV light when it comes in contact with acid phosphatase. Also, 4-methyl umbelliferyl phosphate (MUP) fluoresces under UV light when it comes in contact with acid phosphatase.